Non-myeloablative hematopoietic cell transplantation (HCT) followed by delayed donor leukocyte infusion (DLI) is a promising immunotherapeutic approach to treat hematologic malignancies including leukemias and lymphomas. Major limitations of this approach, however, include the risks of graft failure, graft-versus-host disease, (GVHD) and infection. MGH MHC-inbred miniature swine provide a pre-clinical model for studies of transplantation biology with responses to HCT resembling those of humans. Preliminary data suggest that a novel, minimally myelosuppressive preparative regimen leads to stable multilineage chimerism following highdose haploidentical HCT, without causing GVHD. In this proposal we aim to 1) determine the immunological mechanisms involved in controlling hbst-versus-graft (HVG) and GVH responses that allow engraftment without GVHD across MHC barriers in this model. These studies will be performed in close collaboration with Projects 1 and 2 to extend the mechanistic studies in rodent models. We will then 2) optimize the swine model to facilitate translation of this protocol to the clinic and analyze the importance of specific genetic disparities (haploidentical class I and II, class I only, class II only) on engraftment, GVHD and the effects of subsequent DLI. In collaboration with Project 4, we will test novel strategies to improve stem cell harvests following cytokine mobilization through parathyroid hormone (PTH) stimulation. Increased numbers of stem cells in the leukapheresis product following PTH stimulation and cytokine mobilization may enable stable engraftment using lower doses of cells more easily attainable in the clinic. It is hoped that results of these studies will permit the development of tailored approaches to the use of DLI in chimeric patients depending on their HLA disparities from the donor. In addition, we plan to 3) further develop the swine model to allow direct assessment of graft-versus-tumor effects of HCT and DLI. For this purpose, we will establish tumor cell lines derived from inbred miniature swine and adapt these tumor lines for in vivo growth in pigs. With the recent availability of histocompatible miniature swine, and tumor lines derived from these highly inbred animals, we have the unique opportunity to develop transplantable tumors in a preclinical large animal model. These studies could provide a foundation for future immunotherapeutic approaches for the treatment of hematological malignancies that may be translated toward treatment of human disease.